The Experts below are selected from a list of 264 Experts worldwide ranked by ideXlab platform
Xia Cai - One of the best experts on this subject based on the ideXlab platform.
-
Lysosome and proteasome pathways are distributed in Laticifers of Euphorbia helioscopia L.
Physiologia plantarum, 2018Co-Authors: Xiaoai Fang, Meng Wang, Qing Zhang, Yue Zhang, Bofei Wei, Yan Miao, Lanting Tian, Xia CaiAbstract:At present, the lysosome pathway (LP) and proteasome pathway (PP) are known as major clearance systems in eukaryotic cells. The Laticifer, a secretory tissue, degrades some cytoplasm during development. In this study, we investigated the distribution of LP and PP in non-articulated Laticifers of Euphorbia helioscopia L. Electron microscopy revealed that, plastids, mitochondria and some cyotsol were degraded in the late development Laticifers, where there were numerous vesicles originated from dicytosomes. Accordingly, some key proteins in LP and PP were detected in E. helioscopia latex using isobaric tags for relative and absolute quantitation (iTRAQ) proteomics. Further immunohistochemistry analysis revealed that the clathrin heavy chain (CHC) belonging to LP and the ubiquitin-mediated proteasome degradation increases gradually as the Laticifer develops. Immuno-electron microscopy revealed that the cysteine protease, CHC and AP-2 complex subunit beta-1 belonging to LP were mainly distributed in vesicles deriving from dicytosomes, which we called lysosome-like vesicles. Ubiquitin was widely distributed in the cytosol, and proteasome activity was significantly reduced when various concentrations of the inhibitor MG132 were added to the latex total protein. We hypothesize that LP and PP are distributed in E. helioscopia Laticifers; and it was speculated that LP and PP might be involved in the degradation of organelles and some cytoplasmic matrix in E. helioscopia Laticifers.
-
Detection of autophagy processes during the development of nonarticulated Laticifers in Euphorbia kansui Liou.
Planta, 2017Co-Authors: Qing Zhang, Dou Wang, Meng Wang, Hao Zhang, Xiaoai Fang, Xia CaiAbstract:Main conclusion Autophagy is involved in cytoplasmic degradation through directly engulfing cytosol and organelles by autophagosomes and then fusing with lysosome-like vesicles during the development of nonarticulated Laticifers in Euphorbia kansui Liou. Autophagy has been reported to play an important role in a wide range of eukaryotic organisms during responses to various abiotic and biotic stresses. However, until recently, the functions of autophagy in normal plant differentiation and development were still in their infancy. Nonarticulated Laticifers, a type of secretory tissue in plants, undergo the degradation of cytosol and organelles during their development. However, little evidence of autophagy in Laticifer differentiation has been provided. In the present study, using anti-ATG8 antibody-Alexa Fluor 488, Lyso-Tracker Red (LTR) and monodansylcadaverine (MDC) as markers for detecting autophagosomes, as well as autophagy-related structures, we observed that the green fluorescence of ATG8a largely colocalized with the red fluorescence of LTR and purple fluorescence of MDC and the quantity of autophagosomes experienced a trend from less to more to less during Laticifer development. Additionally, we described the autophagy process during the development of nonarticulated Laticifers in Euphorbia kansui Liou at the ultrastructural level in detail. In addition, further immunogold TEM studies also verified the presence of autophagosomes, autolysosomes and lysosome-like structures in Laticifers. Taken together, these results suggest that autophagy contributes to the development of the nonarticulated Laticifers in E. kansui Liou and that autophagosomes fuse with lysosome-like structures for degradation. These results will lay an important foundation for further studies on Laticifer regulation.
-
Identification and cytochemical immunolocalization of acetyl-CoA acetyltransferase involved in the terpenoid mevalonate pathway in Euphorbia helioscopia Laticifers
Botanical Studies, 2017Co-Authors: Meng Wang, Dou Wang, Jia Chai, Qing Zhang, Yong Peng, Xia CaiAbstract:BackgroundTerpenoids, the largest class of natural products in the plant kingdom, have been widely used in medicine. The precursors of terpenoids, isoprene phosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), were synthesized from a mevalonate (MVA) pathway and a 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway respectively. The acetyl-CoA acetyltransferase (AACT) is the initial enzyme in MVA pathway and is considered presently to be essential for terpenoid backbone biosynthesis. The basic research on cytochemistry of terpenoid metabolic enzymes is important for understanding the mechanisms underlying major metabolic processes. However, compartmentalization of AACT in plants is in controversy. Euphorbia helioscopia L. containing Laticifers in the whole plant is a famous ancient folk medicine for tumor treatment, and the terpenoid is an active ingredient. Furthermore, the Laticifer cell is the main synthesizing and storing site for terpenoids.ResultsThe gene of AACT was cloned successfully from E. helioscopia, and named as EhAACT. The EhAACT expression has no significant difference among roots, stems and leaves. However, compared with the roots and stems, the EhAACT expression level is slightly higher in leaves. In addition, EhAACT recombinant protein was expressed by procaryotic expression system and anti-EhAACT antibody was prepared, the molecular weight is about 43 kDa. Western blotting results illustrated that the EhAACT antibodies specifically recognized the endogenous proteins in E. helioscopia Laticifers. At last, the subcellular localization of EhAACT in E. helioscopia Laticifers was observed by using colloidal gold immune-electron microscopy. EhAACT was found to mainly distribute in the endoplasmic reticulum (ER), vacuoles originated from ER and cytosol aound vacuoles originated from ER.ConclusionsAs a result, we speculated that in E. helioscopia Laticifers, EhAACT located in cytosol would be transferred to small vacuoles dilated from ER, and the precursors of terpenoids were synthesized in these small vacuoles, then terpenoids were further synthesized into latex particles. This result would provide theoretical basis for regulating and controlling of terpenoid biosynthesis in Laticifers.
Jarunya Narangajavana - One of the best experts on this subject based on the ideXlab platform.
-
Comparative proteomic analysis of differentially expressed proteins related to phloem and xylem development in rubber tree (Hevea brasiliensis)
Trees, 2020Co-Authors: Piyawadee Prasongsansuk, Kanlaya Nirapathpongporn, Unchera Viboonjun, Panida Kongsawadworakul, Tipparat Thiangtrongjit, Onrapak Reamtong, Jarunya NarangajavanaAbstract:Key message The proteomic analysis of vascular tissues in rubber tree reveals differentially expressed proteins related to Laticifer differentiation in mature phloem and secondary cell wall formation in mature xylem for latex/wood-yield improvement. Abstract Rubber tree ( Hevea brasiliensis ) is a latex-producing plant that has worldwide economic importance for the rubber and wood industries. Natural rubber latex is produced from Laticiferous vessels (Laticifers) located in the secondary phloem tissue of rubber trees. Improving latex production from rubber tree clones by studying the molecular genetics of vascular development has received much attention, but less information has been obtained from proteomic approaches. This study performed a comparative proteomic analysis of the vascular tissues in high latex-yield and high wood-yield rubber tree clones. One primary vascular tissue (newly developed stem) and two secondary vascular tissues (mature phloem/Laticifers and mature xylem/wood) were investigated by qualitative/quantitative proteomic analysis using GeLC-MS/MS. The differentially expressed proteins (DEPs) in the specific vascular tissues were analyzed, and the protein functions in the biological processes related to vascular development and to the specific characteristics of each clone were clarified. The predicted protein–protein interaction network and GO annotation revealed DEPs related to photosynthesis, carbohydrate metabolism, energy production and jasmonic acid-responsive proteins involved in positive regulation of Laticifer differentiation in mature phloem of the high latex-yield clone, while DEPs related to cell cytoskeleton maintenance, secondary cell wall components and auxin-, brassinosteroid-, abscisic acid-responsive proteins involved in xylem differentiation were abundant in mature xylem of the high wood-yield clone. This is the first report to demonstrate the correlation and functions of DEPs in phloem/Laticifers and xylem/wood differentiation in rubber trees. The regulation of vascular development could be useful in improvement of latex and wood yields from rubber trees.
-
Comparative proteomic analysis of differentially expressed proteins related to phloem and xylem development in rubber tree (Hevea brasiliensis)
Trees, 2020Co-Authors: Piyawadee Prasongsansuk, Kanlaya Nirapathpongporn, Unchera Viboonjun, Panida Kongsawadworakul, Tipparat Thiangtrongjit, Onrapak Reamtong, Jarunya NarangajavanaAbstract:Key message The proteomic analysis of vascular tissues in rubber tree reveals differentially expressed proteins related to Laticifer differentiation in mature phloem and secondary cell wall formation in mature xylem for latex/wood-yield improvement. Abstract Rubber tree ( Hevea brasiliensis ) is a latex-producing plant that has worldwide economic importance for the rubber and wood industries. Natural rubber latex is produced from Laticiferous vessels (Laticifers) located in the secondary phloem tissue of rubber trees. Improving latex production from rubber tree clones by studying the molecular genetics of vascular development has received much attention, but less information has been obtained from proteomic approaches. This study performed a comparative proteomic analysis of the vascular tissues in high latex-yield and high wood-yield rubber tree clones. One primary vascular tissue (newly developed stem) and two secondary vascular tissues (mature phloem/Laticifers and mature xylem/wood) were investigated by qualitative/quantitative proteomic analysis using GeLC-MS/MS. The differentially expressed proteins (DEPs) in the specific vascular tissues were analyzed, and the protein functions in the biological processes related to vascular development and to the specific characteristics of each clone were clarified. The predicted protein–protein interaction network and GO annotation revealed DEPs related to photosynthesis, carbohydrate metabolism, energy production and jasmonic acid-responsive proteins involved in positive regulation of Laticifer differentiation in mature phloem of the high latex-yield clone, while DEPs related to cell cytoskeleton maintenance, secondary cell wall components and auxin-, brassinosteroid-, abscisic acid-responsive proteins involved in xylem differentiation were abundant in mature xylem of the high wood-yield clone. This is the first report to demonstrate the correlation and functions of DEPs in phloem/Laticifers and xylem/wood differentiation in rubber trees. The regulation of vascular development could be useful in improvement of latex and wood yields from rubber trees.
-
unraveling vascular development related genes in Laticifer containing tissue of rubber tree by high throughput transcriptome sequencing
Current Plant Biology, 2019Co-Authors: Pakatorn Saelim, Kanlaya Nirapathpongporn, Unchera Viboonjun, Panida Kongsawadworakul, Chaiwat Naktang, Thippawan Yoocha, Sithichoke Tangphatsornruang, Jarunya NarangajavanaAbstract:Abstract About 90% of natural rubber is obtained from tapping of the rubber tree (Hevea brasiliensis) for latex, which is circulated in the Laticifers. Nowadays, the world supply of natural rubber is not sufficient for global demand, thus the increased latex yield was significantly underlined. In addition, the demand for wood (as a xylem part) from rubber tree was also increased as renewable resource for various applications. Laticifers are found in the secondary phloem containing tissue of rubber tree trunk-inner soft bark. The number of Laticifers varies in consistent with latex yield and in responses to jasmonic acid level. This present study was committed to comparative transcriptome analysis in Laticifers containing mature phloem, mature xylem and newly developed stem tissues of high latex-yield clone (RRIT251, with more Laticifers) and high wood-yield clone (RRIT402, with less Laticifers) of rubber tree to classify the genes and pathways involved with phloem (with Laticifers) and xylem cell differentiation. There were 49, 54, 46 and 50 of vascular development-related genes in primary and secondary tissues of phloem and xylem, respectively. Differentially expressed genes in jasmonic acid signaling pathway was established with their highest expression in phloem tissue with Laticifer cells of RRIT251, while genes in auxin signaling and secondary cell wall biosynthetic pathways were up-regulated in xylem tissue of RRIT402 for high wood yield. Promoter analysis of candidate-differentially expressed genes suggested the related pathway and putative regulatory elements for gene regulation. This genome-wide exploration of vascular development-related genes unraveled a largely unknown gap of this special vascular development containing Laticifers in rubber tree.
-
opposite physiological effects upon jasmonic acid and brassinosteroid treatment on Laticifer proliferation and co occurrence of differential expression of genes involved in vascular development in rubber tree
Physiology and Molecular Biology of Plants, 2019Co-Authors: Poochita Arreewichit, Pakatorn Saelim, Kanlaya Nirapathpongporn, Unchera Viboonjun, Panida Kongsawadworakul, Jarunya NarangajavanaAbstract:During growth of woody plant-trunk, the secondary meristem functions in giving rise the xylem and phloem. Rubber tree (Hevea brasiliensis Muell. Arg.), in addition, contains Laticifers (latex producing vessels) in the vicinity of phloem. Insights into regulatory mechanisms of gene networks underlying Laticifer proliferation in rubber tree has remained very limited. The candidate vascular development-related genes were selected to investigate for expression profile in phloem and xylem tissues of high latex yield- and high wood yield-clones of rubber tree. The differential gene expression between the mature branch-xylem and -phloem tissues was clearly observed. The cis-regulatory motif analysis revealed the existent of putative jasmonic acid (JA)- and brassinosteroid (BR)-responsive regulatory motifs in promoter regions of these genes, and consequently the effect of exogenous application of JA, BR or their respective signaling inhibitors, on the formation of Laticifers in rubber tree was demonstrated. Interestingly, the Laticifer numbers were significantly increased in JA-treatment, correlated with up-regulation of phloem development-related genes in both rubber tree clones. On the contrary, the Laticifers were decreased in BR-treatment accompanying by up-regulation of xylem development-related genes, especially in high wood yield-rubber tree clone. BR-inhibitor treatment also enhanced Laticifer numbers, while JA-inhibitor suppressed Laticifer differentiation. Taken together, this study unveils the molecular interplay between JA/BR on vascular development in rubber tree and how this impacts the appearance of Laticifers in this plant. This process is vital for a better understanding on Laticifer differentiation and its impact in the manipulation of wood and latex yield in rubber tree improvement program.
-
Involvement of HbPIP2;1 and HbTIP1;1 Aquaporins in Ethylene Stimulation of Latex Yield through Regulation of Water Exchanges between Inner Liber and Latex Cells in Hevea brasiliensis
Plant Physiology, 2009Co-Authors: Kessarin Tungngoen, Unchera Viboonjun, Panida Kongsawadworakul, Jarunya Narangajavana, Maki Katsuhara, Nicole Brunel, Soulaiman Sakr, Hervé ChrestinAbstract:Natural rubber is synthesized in specialized articulated cells (Laticifers) located in the inner liber of Hevea brasiliensis. Upon bark tapping, the Laticifer cytoplasm (latex) is expelled due to liber tissue turgor pressure. In mature virgin (untapped) trees, short-term kinetic studies confirmed that ethylene, the rubber yield stimulant used worldwide, increased latex yield, with a concomitant decrease in latex total solid content, probably through water influx in the Laticifers. As the mature Laticifers are devoid of plasmodesmata, the rapid water exchanges with surrounding liber cells probably occur via the aquaporin pathway. Two full-length aquaporin cDNAs (HbPIP2;1 and HbTIP1;1, for plasma membrane intrinsic protein and tonoplast intrinsic protein, respectively) were cloned and characterized. The higher efficiency of HbPIP2;1 than HbTIP1;1 in increasing plasmalemma water conductance was verified in Xenopus laevis oocytes. HbPIP2;1 was insensitive to HgCl(2). In situ hybridization demonstrated that HbPIP2;1 was expressed in all liber tissues in the young stem, including the Laticifers. HbPIP2;1 was up-regulated in both liber tissues and Laticifers, whereas HbTIP1;1 was down-regulated in liber tissues but up-regulated in Laticifers in response to bark Ethrel treatment. Ethylene-induced HbPIP2;1 up-regulation was confirmed by western-blot analysis. The promoter sequences of both genes were cloned and found to harbor, among many others, ethylene-responsive and other chemical-responsive (auxin, copper, and sulfur) elements known to increase latex yield. Increase in latex yield in response to ethylene was emphasized to be linked with water circulation between the Laticifers and their surrounding tissues as well as with the probable maintenance of liber tissue turgor, which together favor prolongation of latex flow.
Dirk Prufer - One of the best experts on this subject based on the ideXlab platform.
-
comparative proteome and metabolome analyses of latex exuding and non exuding taraxacum koksaghyz roots provide insights into Laticifer biology
Journal of Experimental Botany, 2020Co-Authors: Vincent Alexander Benninghaus, Dirk Prufer, Nicole Van Deenen, Boje Muller, Kaiuwe Roelfs, Ines Lassowskat, Iris Finkemeier, Christian Schulze GronoverAbstract:Taraxacum koksaghyz has been identified as one of the most promising alternative rubber crops. Its high-quality rubber is produced in the latex of Laticifers, a specialized cell type that is organized in a network of elongated tubules throughout the entire plant body. In order to gain insights into the physiological role(s) of latex and hence Laticifer biology, we examine the effects of barnase-induced latex RNA degradation on the metabolite and protein compositions in the roots. We established high-quality datasets that enabled precise discrimination between cellular and physiological processes in Laticifers and non-Laticifer cell types of roots at different vegetative stages. We identified numerous latex-specific proteins, including a perilipin-like protein that has not been studied in plants yet. The barnase-expressing plants revealed a phenotype that did not exude latex, which may provide a valuable genetic basis for future studies of plant-environment interactions concerning latex and also help to clarify the evolution and arbitrary distribution of latex throughout the plant kingdom. The overview of temporal changes in composition and protein abundance provided by our data opens the way for a deeper understanding of the molecular interactions, reactions, and network relationships that underlie the different metabolic pathways in the roots of this potential rubber crop.
-
establishment of an ex vivo Laticifer cell suspension culture from taraxacum brevicorniculatum as a production system for cis isoprene
Journal of Molecular Catalysis B-enzymatic, 2014Co-Authors: Janina Post, Wolfgang Eisenreich, Claudia Huber, Richard M Twyman, Dirk Prufer, Christian Schulze GronoverAbstract:Abstract Laticifers are highly specialized plant cells that produce latex enriched with secondary metabolites. The articulated Laticifers of Taraxacum brevicorniculatum synthesize natural rubber, an industrially-valuable composite biopolymer comprising >95% high-molecular-weight (HMW) poly( cis -1,4-isoprene). Here we present a proof-of-concept approach for the cultivation of cell suspension cultures exclusively containing Laticifers. We transformed T. brevicorniculatum plants with a plasmid conferring Laticifer-specific hygromycin resistance. Transgenic callus tissue was used to induce a cell suspension culture under antibiotic selection to favor Laticifer growth. The cultured cells appeared Laticiferous in terms of morphology and expressed Laticifer-specific genes. Confocal laser scanning microscopy revealed intracellular lipid accumulation in vesicle-like structures. Nuclear magnetic resonance and diffusion ordered spectroscopy indicated the presence of mid-length poly( cis -1,4-isoprene) chains but no high HMW natural rubber in the cells. Precursor feeding with mevalonolactone increased the accumulation of poly( cis -1,4-isoprene) by 17-fold, reaching a concentration of 2.7 mg/g dry weight. Our approach could lead to the development of a production platform for the efficient conversion of isopentenyl diphosphate into poly( cis -1,4-isoprene) in an optimized cell suspension culture system. The apparent absence of HMW natural rubber is discussed in terms of our current knowledge of rubber biosynthesis.
-
Laticifer specific cis prenyltransferase silencing affects the rubber triterpene and inulin content of taraxacum brevicorniculatum
Plant Physiology, 2012Co-Authors: Janina Post, Wolfgang Eisenreich, Claudia Huber, Richard M Twyman, Nicole Van Deenen, Julia Fricke, Natalie Kowalski, David Wurbs, Hubert Schaller, Dirk PruferAbstract:Certain Taraxacum species, such as Taraxacum koksaghyz and Taraxacum brevicorniculatum, produce large amounts of high-quality natural rubber in their latex, the milky cytoplasm of specialized cells known as Laticifers. This high-molecular mass biopolymer consists mainly of poly(cis-1,4-isoprene) and is deposited in rubber particles by particle-bound enzymes that carry out the stereospecific condensation of isopentenyl diphosphate units. The polymer configuration suggests that the chain-elongating enzyme (rubber transferase; EC 2.5.1.20) is a cis-prenyltransferase (CPT). Here, we present a comprehensive analysis of transgenic T. brevicorniculatum plants in which the expression of three recently isolated CPTs known to be associated with rubber particles (TbCPT1 to -3) was heavily depleted by Laticifer-specific RNA interference (RNAi). Analysis of the CPT-RNAi plants by nuclear magnetic resonance, size-exclusion chromatography, and gas chromatography-mass spectrometry indicated a significant reduction in rubber biosynthesis and a corresponding 50% increase in the levels of triterpenes and the main storage carbohydrate, inulin. Transmission electron microscopy revealed that the Laticifers in CPT-RNAi plants contained fewer and smaller rubber particles than wild-type Laticifers. We also observed lower activity of hydroxymethylglutaryl-coenzyme A reductase, the key enzyme in the mevalonate pathway, reflecting homeostatic control of the isopentenyl diphosphate pool. To our knowledge, this is the first in planta demonstration of latex-specific CPT activity in rubber biosynthesis.
-
polyphenoloxidase silencing affects latex coagulation in taraxacum species
Plant Physiology, 2009Co-Authors: Daniela Wahler, Richard M Twyman, Christian Schulze Gronover, Carolin Richter, Florence Foucu, Bruno M Moerschbacher, Rainer Fischer, Jost Muth, Dirk PruferAbstract:Latex is the milky sap that is found in many different plants. It is produced by specialized cells known as Laticifers and can comprise a mixture of proteins, carbohydrates, oils, secondary metabolites, and rubber that may help to prevent herbivory and protect wound sites against infection. The wound-induced browning of latex suggests that it contains one or more phenol-oxidizing enzymes. Here, we present a comprehensive analysis of the major latex proteins from two dandelion species, Taraxacum officinale and Taraxacum kok-saghyz, and enzymatic studies showing that polyphenoloxidase (PPO) is responsible for latex browning. Electrophoretic analysis and amino-terminal sequencing of the most abundant proteins in the aqueous latex fraction revealed the presence of three PPO-related proteins generated by the proteolytic cleavage of a single precursor (pre-PPO). The Laticifer-specific pre-PPO protein contains a transit peptide that can target reporter proteins into chloroplasts when constitutively expressed in dandelion protoplasts, perhaps indicating the presence of structures similar to plastids in Laticifers, which lack genuine chloroplasts. Silencing the PPO gene by constitutive RNA interference in transgenic plants reduced PPO activity compared with wild-type controls, allowing T. kok-saghyz RNA interference lines to expel four to five times more latex than controls. Latex fluidity analysis in silenced plants showed a strong correlation between residual PPO activity and the coagulation rate, indicating that Laticifer-specific PPO plays a major role in latex coagulation and wound sealing in dandelions. In contrast, very little PPO activity is found in the latex of the rubber tree Hevea brasiliensis, suggesting functional divergence of latex proteins during plant evolution.
Meng Wang - One of the best experts on this subject based on the ideXlab platform.
-
Lysosome and proteasome pathways are distributed in Laticifers of Euphorbia helioscopia L.
Physiologia plantarum, 2018Co-Authors: Xiaoai Fang, Meng Wang, Qing Zhang, Yue Zhang, Bofei Wei, Yan Miao, Lanting Tian, Xia CaiAbstract:At present, the lysosome pathway (LP) and proteasome pathway (PP) are known as major clearance systems in eukaryotic cells. The Laticifer, a secretory tissue, degrades some cytoplasm during development. In this study, we investigated the distribution of LP and PP in non-articulated Laticifers of Euphorbia helioscopia L. Electron microscopy revealed that, plastids, mitochondria and some cyotsol were degraded in the late development Laticifers, where there were numerous vesicles originated from dicytosomes. Accordingly, some key proteins in LP and PP were detected in E. helioscopia latex using isobaric tags for relative and absolute quantitation (iTRAQ) proteomics. Further immunohistochemistry analysis revealed that the clathrin heavy chain (CHC) belonging to LP and the ubiquitin-mediated proteasome degradation increases gradually as the Laticifer develops. Immuno-electron microscopy revealed that the cysteine protease, CHC and AP-2 complex subunit beta-1 belonging to LP were mainly distributed in vesicles deriving from dicytosomes, which we called lysosome-like vesicles. Ubiquitin was widely distributed in the cytosol, and proteasome activity was significantly reduced when various concentrations of the inhibitor MG132 were added to the latex total protein. We hypothesize that LP and PP are distributed in E. helioscopia Laticifers; and it was speculated that LP and PP might be involved in the degradation of organelles and some cytoplasmic matrix in E. helioscopia Laticifers.
-
Detection of autophagy processes during the development of nonarticulated Laticifers in Euphorbia kansui Liou.
Planta, 2017Co-Authors: Qing Zhang, Dou Wang, Meng Wang, Hao Zhang, Xiaoai Fang, Xia CaiAbstract:Main conclusion Autophagy is involved in cytoplasmic degradation through directly engulfing cytosol and organelles by autophagosomes and then fusing with lysosome-like vesicles during the development of nonarticulated Laticifers in Euphorbia kansui Liou. Autophagy has been reported to play an important role in a wide range of eukaryotic organisms during responses to various abiotic and biotic stresses. However, until recently, the functions of autophagy in normal plant differentiation and development were still in their infancy. Nonarticulated Laticifers, a type of secretory tissue in plants, undergo the degradation of cytosol and organelles during their development. However, little evidence of autophagy in Laticifer differentiation has been provided. In the present study, using anti-ATG8 antibody-Alexa Fluor 488, Lyso-Tracker Red (LTR) and monodansylcadaverine (MDC) as markers for detecting autophagosomes, as well as autophagy-related structures, we observed that the green fluorescence of ATG8a largely colocalized with the red fluorescence of LTR and purple fluorescence of MDC and the quantity of autophagosomes experienced a trend from less to more to less during Laticifer development. Additionally, we described the autophagy process during the development of nonarticulated Laticifers in Euphorbia kansui Liou at the ultrastructural level in detail. In addition, further immunogold TEM studies also verified the presence of autophagosomes, autolysosomes and lysosome-like structures in Laticifers. Taken together, these results suggest that autophagy contributes to the development of the nonarticulated Laticifers in E. kansui Liou and that autophagosomes fuse with lysosome-like structures for degradation. These results will lay an important foundation for further studies on Laticifer regulation.
-
Identification and cytochemical immunolocalization of acetyl-CoA acetyltransferase involved in the terpenoid mevalonate pathway in Euphorbia helioscopia Laticifers
Botanical Studies, 2017Co-Authors: Meng Wang, Dou Wang, Jia Chai, Qing Zhang, Yong Peng, Xia CaiAbstract:BackgroundTerpenoids, the largest class of natural products in the plant kingdom, have been widely used in medicine. The precursors of terpenoids, isoprene phosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), were synthesized from a mevalonate (MVA) pathway and a 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway respectively. The acetyl-CoA acetyltransferase (AACT) is the initial enzyme in MVA pathway and is considered presently to be essential for terpenoid backbone biosynthesis. The basic research on cytochemistry of terpenoid metabolic enzymes is important for understanding the mechanisms underlying major metabolic processes. However, compartmentalization of AACT in plants is in controversy. Euphorbia helioscopia L. containing Laticifers in the whole plant is a famous ancient folk medicine for tumor treatment, and the terpenoid is an active ingredient. Furthermore, the Laticifer cell is the main synthesizing and storing site for terpenoids.ResultsThe gene of AACT was cloned successfully from E. helioscopia, and named as EhAACT. The EhAACT expression has no significant difference among roots, stems and leaves. However, compared with the roots and stems, the EhAACT expression level is slightly higher in leaves. In addition, EhAACT recombinant protein was expressed by procaryotic expression system and anti-EhAACT antibody was prepared, the molecular weight is about 43 kDa. Western blotting results illustrated that the EhAACT antibodies specifically recognized the endogenous proteins in E. helioscopia Laticifers. At last, the subcellular localization of EhAACT in E. helioscopia Laticifers was observed by using colloidal gold immune-electron microscopy. EhAACT was found to mainly distribute in the endoplasmic reticulum (ER), vacuoles originated from ER and cytosol aound vacuoles originated from ER.ConclusionsAs a result, we speculated that in E. helioscopia Laticifers, EhAACT located in cytosol would be transferred to small vacuoles dilated from ER, and the precursors of terpenoids were synthesized in these small vacuoles, then terpenoids were further synthesized into latex particles. This result would provide theoretical basis for regulating and controlling of terpenoid biosynthesis in Laticifers.
-
Identification and cytochemical immunolocalization of acetyl-CoA acetyltransferase involved in the terpenoid mevalonate pathway in Euphorbia helioscopia Laticifers.
Botanical Studies, 2017Co-Authors: Meng Wang, Dou Wang, Jia Chai, Qing Zhang, Yong PengAbstract:Terpenoids, the largest class of natural products in the plant kingdom, have been widely used in medicine. The precursors of terpenoids, isoprene phosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), were synthesized from a mevalonate (MVA) pathway and a 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway respectively. The acetyl-CoA acetyltransferase (AACT) is the initial enzyme in MVA pathway and is considered presently to be essential for terpenoid backbone biosynthesis. The basic research on cytochemistry of terpenoid metabolic enzymes is important for understanding the mechanisms underlying major metabolic processes. However, compartmentalization of AACT in plants is in controversy. Euphorbia helioscopia L. containing Laticifers in the whole plant is a famous ancient folk medicine for tumor treatment, and the terpenoid is an active ingredient. Furthermore, the Laticifer cell is the main synthesizing and storing site for terpenoids. The gene of AACT was cloned successfully from E. helioscopia, and named as EhAACT. The EhAACT expression has no significant difference among roots, stems and leaves. However, compared with the roots and stems, the EhAACT expression level is slightly higher in leaves. In addition, EhAACT recombinant protein was expressed by procaryotic expression system and anti-EhAACT antibody was prepared, the molecular weight is about 43 kDa. Western blotting results illustrated that the EhAACT antibodies specifically recognized the endogenous proteins in E. helioscopia Laticifers. At last, the subcellular localization of EhAACT in E. helioscopia Laticifers was observed by using colloidal gold immune-electron microscopy. EhAACT was found to mainly distribute in the endoplasmic reticulum (ER), vacuoles originated from ER and cytosol aound vacuoles originated from ER. As a result, we speculated that in E. helioscopia Laticifers, EhAACT located in cytosol would be transferred to small vacuoles dilated from ER, and the precursors of terpenoids were synthesized in these small vacuoles, then terpenoids were further synthesized into latex particles. This result would provide theoretical basis for regulating and controlling of terpenoid biosynthesis in Laticifers.
Qing Zhang - One of the best experts on this subject based on the ideXlab platform.
-
Lysosome and proteasome pathways are distributed in Laticifers of Euphorbia helioscopia L.
Physiologia plantarum, 2018Co-Authors: Xiaoai Fang, Meng Wang, Qing Zhang, Yue Zhang, Bofei Wei, Yan Miao, Lanting Tian, Xia CaiAbstract:At present, the lysosome pathway (LP) and proteasome pathway (PP) are known as major clearance systems in eukaryotic cells. The Laticifer, a secretory tissue, degrades some cytoplasm during development. In this study, we investigated the distribution of LP and PP in non-articulated Laticifers of Euphorbia helioscopia L. Electron microscopy revealed that, plastids, mitochondria and some cyotsol were degraded in the late development Laticifers, where there were numerous vesicles originated from dicytosomes. Accordingly, some key proteins in LP and PP were detected in E. helioscopia latex using isobaric tags for relative and absolute quantitation (iTRAQ) proteomics. Further immunohistochemistry analysis revealed that the clathrin heavy chain (CHC) belonging to LP and the ubiquitin-mediated proteasome degradation increases gradually as the Laticifer develops. Immuno-electron microscopy revealed that the cysteine protease, CHC and AP-2 complex subunit beta-1 belonging to LP were mainly distributed in vesicles deriving from dicytosomes, which we called lysosome-like vesicles. Ubiquitin was widely distributed in the cytosol, and proteasome activity was significantly reduced when various concentrations of the inhibitor MG132 were added to the latex total protein. We hypothesize that LP and PP are distributed in E. helioscopia Laticifers; and it was speculated that LP and PP might be involved in the degradation of organelles and some cytoplasmic matrix in E. helioscopia Laticifers.
-
Detection of autophagy processes during the development of nonarticulated Laticifers in Euphorbia kansui Liou.
Planta, 2017Co-Authors: Qing Zhang, Dou Wang, Meng Wang, Hao Zhang, Xiaoai Fang, Xia CaiAbstract:Main conclusion Autophagy is involved in cytoplasmic degradation through directly engulfing cytosol and organelles by autophagosomes and then fusing with lysosome-like vesicles during the development of nonarticulated Laticifers in Euphorbia kansui Liou. Autophagy has been reported to play an important role in a wide range of eukaryotic organisms during responses to various abiotic and biotic stresses. However, until recently, the functions of autophagy in normal plant differentiation and development were still in their infancy. Nonarticulated Laticifers, a type of secretory tissue in plants, undergo the degradation of cytosol and organelles during their development. However, little evidence of autophagy in Laticifer differentiation has been provided. In the present study, using anti-ATG8 antibody-Alexa Fluor 488, Lyso-Tracker Red (LTR) and monodansylcadaverine (MDC) as markers for detecting autophagosomes, as well as autophagy-related structures, we observed that the green fluorescence of ATG8a largely colocalized with the red fluorescence of LTR and purple fluorescence of MDC and the quantity of autophagosomes experienced a trend from less to more to less during Laticifer development. Additionally, we described the autophagy process during the development of nonarticulated Laticifers in Euphorbia kansui Liou at the ultrastructural level in detail. In addition, further immunogold TEM studies also verified the presence of autophagosomes, autolysosomes and lysosome-like structures in Laticifers. Taken together, these results suggest that autophagy contributes to the development of the nonarticulated Laticifers in E. kansui Liou and that autophagosomes fuse with lysosome-like structures for degradation. These results will lay an important foundation for further studies on Laticifer regulation.
-
Identification and cytochemical immunolocalization of acetyl-CoA acetyltransferase involved in the terpenoid mevalonate pathway in Euphorbia helioscopia Laticifers
Botanical Studies, 2017Co-Authors: Meng Wang, Dou Wang, Jia Chai, Qing Zhang, Yong Peng, Xia CaiAbstract:BackgroundTerpenoids, the largest class of natural products in the plant kingdom, have been widely used in medicine. The precursors of terpenoids, isoprene phosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), were synthesized from a mevalonate (MVA) pathway and a 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway respectively. The acetyl-CoA acetyltransferase (AACT) is the initial enzyme in MVA pathway and is considered presently to be essential for terpenoid backbone biosynthesis. The basic research on cytochemistry of terpenoid metabolic enzymes is important for understanding the mechanisms underlying major metabolic processes. However, compartmentalization of AACT in plants is in controversy. Euphorbia helioscopia L. containing Laticifers in the whole plant is a famous ancient folk medicine for tumor treatment, and the terpenoid is an active ingredient. Furthermore, the Laticifer cell is the main synthesizing and storing site for terpenoids.ResultsThe gene of AACT was cloned successfully from E. helioscopia, and named as EhAACT. The EhAACT expression has no significant difference among roots, stems and leaves. However, compared with the roots and stems, the EhAACT expression level is slightly higher in leaves. In addition, EhAACT recombinant protein was expressed by procaryotic expression system and anti-EhAACT antibody was prepared, the molecular weight is about 43 kDa. Western blotting results illustrated that the EhAACT antibodies specifically recognized the endogenous proteins in E. helioscopia Laticifers. At last, the subcellular localization of EhAACT in E. helioscopia Laticifers was observed by using colloidal gold immune-electron microscopy. EhAACT was found to mainly distribute in the endoplasmic reticulum (ER), vacuoles originated from ER and cytosol aound vacuoles originated from ER.ConclusionsAs a result, we speculated that in E. helioscopia Laticifers, EhAACT located in cytosol would be transferred to small vacuoles dilated from ER, and the precursors of terpenoids were synthesized in these small vacuoles, then terpenoids were further synthesized into latex particles. This result would provide theoretical basis for regulating and controlling of terpenoid biosynthesis in Laticifers.
-
Identification and cytochemical immunolocalization of acetyl-CoA acetyltransferase involved in the terpenoid mevalonate pathway in Euphorbia helioscopia Laticifers.
Botanical Studies, 2017Co-Authors: Meng Wang, Dou Wang, Jia Chai, Qing Zhang, Yong PengAbstract:Terpenoids, the largest class of natural products in the plant kingdom, have been widely used in medicine. The precursors of terpenoids, isoprene phosphate (IPP) and dimethylallyl pyrophosphate (DMAPP), were synthesized from a mevalonate (MVA) pathway and a 2-C-methyl-d-erythritol-4-phosphate (MEP) pathway respectively. The acetyl-CoA acetyltransferase (AACT) is the initial enzyme in MVA pathway and is considered presently to be essential for terpenoid backbone biosynthesis. The basic research on cytochemistry of terpenoid metabolic enzymes is important for understanding the mechanisms underlying major metabolic processes. However, compartmentalization of AACT in plants is in controversy. Euphorbia helioscopia L. containing Laticifers in the whole plant is a famous ancient folk medicine for tumor treatment, and the terpenoid is an active ingredient. Furthermore, the Laticifer cell is the main synthesizing and storing site for terpenoids. The gene of AACT was cloned successfully from E. helioscopia, and named as EhAACT. The EhAACT expression has no significant difference among roots, stems and leaves. However, compared with the roots and stems, the EhAACT expression level is slightly higher in leaves. In addition, EhAACT recombinant protein was expressed by procaryotic expression system and anti-EhAACT antibody was prepared, the molecular weight is about 43 kDa. Western blotting results illustrated that the EhAACT antibodies specifically recognized the endogenous proteins in E. helioscopia Laticifers. At last, the subcellular localization of EhAACT in E. helioscopia Laticifers was observed by using colloidal gold immune-electron microscopy. EhAACT was found to mainly distribute in the endoplasmic reticulum (ER), vacuoles originated from ER and cytosol aound vacuoles originated from ER. As a result, we speculated that in E. helioscopia Laticifers, EhAACT located in cytosol would be transferred to small vacuoles dilated from ER, and the precursors of terpenoids were synthesized in these small vacuoles, then terpenoids were further synthesized into latex particles. This result would provide theoretical basis for regulating and controlling of terpenoid biosynthesis in Laticifers.